The invention relates to a pharmaceutical composition especially to be used as a vaccine.
Vaccines are a very successful, yet cost saving medical intervention. Several catastrophic illnesses including small pox and poliomyelitis have been, due to intense vaccination programmes, eliminated from the face of this earth or are on the brink of extinction (Nossal, Nat Med 4, (1998), 475-476). In fact, vaccines can save more lives (and money) than any other medical intervention. Although this notion is valid for a whole panel of diseases including tuberculosis, diphteria, pertussis, measles and tetanus, there are no effective vaccines for numerous ailments including most viral infections, such as AIDS, and other illnesses including malaria, or even cancer. In addition, the rapid emergence of antibiotic resistant bacteria and microorganisms calls for alternative treatments with vaccines being a logical choice. Finally, the great need for vaccines is also illustrated by the fact that infectious diseases, rather than cardiovascular disorders or cancer or injuries remain the largest cause of death and disability in the world (Bloom et al, Nat Med 4, (1998), 480-484).
The main problem in the field of vaccines is that traditional vaccines (and/or the immune modulating compounds contained within these preparations) are designed to induce high levels of antibodies (Harlow et al, Cold Spring Harbor: Cold Spring Harbor Laboratory, (1988)). Unfortunately, antibodies on their own are not effective in preventing many diseases including most illnesses caused by viruses, intracellular bacteria, or certain parasites. Examples are pathogens such as the above-mentioned HIV virus or Plasmodium spec. in case of malaria. In addition, these vaccines likely will not be effective in cancer. In numerous experimental systems it has been shown that the cellular arm of the immune system, including T cells, rather than the humoral arm, is important for these indications. Therefore, novel, innovative technologies to overcome the limitations of conventional vaccines are needed. The focus must be on technologies that reliably induce the cellular immune system, including antigen specific T cells, which recognize molecules expressed on pathogen infected cells. Ideally, vaccines are designed that induce both T cells distinguishing diseased, and/or infected cells from normal cells and, simultaneously, antibodies secreted by B cells recognizing pathogens in extracellular compartments.
Commonly, vaccines are administered as a combination of pathogen-derived antigens together with compounds that induce or enhance immune responses against these antigens (these compounds are usually termed adjuvants). Examples of antigens are whole organisms such as inactivated or attenuated viruses or bacteria, fungi, protozoa or even cancer cells. Antigens may also consist of subfractions of theses organism/tissues, proteins or, in their most simple form, peptides. Antigens can also be recognized by the immune system in form of glycosylated proteins or peptides and may also be or contain polysaccharides or lipids. Short peptides can be used since for example cytotoxic T cells recognize antigens in form of short usually 8-11 amino acids long peptides in conjunction with major histocompatibility complex (MHC) (Rammensee et al., Immunogenetics 41, (1995), 178-228). B cells recognize longer peptides starting at around 15 amino acids (Harlow et al, Cold Spring Harbor: Cold Spring Harbor Laboratory, (1988)). By contrast to T cell epitopes the three dimensional structure of B cell antigens may also be important for recognition by antibodies. In order to obtain sustained, antigen-specific immune responses, adjuvants need to trigger immune cascades that involve all cells of the immune system necessary. Primarily, said adjuvants are acting, but are not restricted in their mode of action, on so-called antigen presenting cells (APCs). These cells usually first encounter the antigen(s) followed by presentation of processed or unmodified antigen to immune effector cells. Intermediate cell types may also be involved. Only effector cells with the appropriate specificity are activated in a productive immune response. The adjuvant may also locally retain antigens and co-injected other factors. In addition the adjuvant may act as a chemoattractant for other immune cells or may act locally and/or systemically as a stimulating agent for the immune system.
Human growth hormone (HGH) is a pituitary-derived factor primarily described for its ability to promote growth acceleration (reviewed in Neely et al, Annu Rev Med 45, (1994), 407-420). The first patient was treated with growth hormone obtained from pituitary extracts as early as 1958. Recombinant HGH is available for roughly 15 years now and has been used extensively in the clinic. Side effects of recombinant HGH are rare. Efficacy of recombinant HGH preparations has been demonstrated in a wide spectrum of diseases including Turner syndrome, idiopathic short stature, growth hormone deficiency and renal failure.
Whilst numerous studies have confirmed the growth promoting effect of HGH, relatively few reports address a possible interaction of this molecule with cells of the immune system. Stephenson and Melling, who showed that HGH greatly enhances the efficacy of a viral vaccine preparation, first demonstrated the usefulness of HGH in a vaccine context (Stephenson et al., J Infect Dis 164, (1991), 188-191). They co-injected HGH with a vaccine for tick-born encephalitis (TBE) virus, an endemic virus transmitted by ticks. In animal experiments HGH potentiated vaccine efficacy and led to the protection of animals after only one injection of the vaccine. The mechanism of how HGH enhanced vaccine efficacy is unclear, but it was speculated that cell mediated immunity played a significant role. There is further, albeit circumstantial, evidence that HGH may indeed induce cellular immune reactions: Mellado et al. demonstrated that if an antigen derived from human immuno deficiency virus (HIV) is applied to mice, they develop a so called Thl type T helper cell response indicative of a cellular immune response (Mellado et al., Vaccine 16, (1998), 1111-1115). Taken together, there is circumstantial evidence that HGH, which is viewed as exemplary for a whole class of primarily neuroactive compounds (see e.g. Levite, PNAS 95 (1998), 12544-12549, Scholzen et al, Exp. Dermetal. 7 (1998), 81-96), may have a positive effect on the immune system, but the mechanisms remain unclear.
Polycationic polymers, for example the polycationic amino acid polymers poly-L-arginine and poly-L-lysine, have been shown to allow very efficient charging of antigen presenting cells (APCs) with antigens in vitro and in vivo (Buschle et al., Gene Ther Mol Biol 1, (1998), 309-321; Buschle et al., Proc Natl Acad Sci USA 94, (1997), 3256-3261; Schmidt et al., Proc Natl Acad Sci USA 94, (1997), 3262-3267). This is thought to be the key event for triggering immune cascades eventually leading to the induction of antigen specific immune effector cells that are able to destroy or neutralize targets. It has been shown previously that a number of polycationic compounds exert effects on immune cells (Buschle et al., Gene Ther Mol Biol 1, (1998), 309-321; Buschle et al., Proc Natl Acad Sci USA 94, (1997), 3256-3261).
Co-injection of a mixture of poly-L-arginine or poly-L-lysine together with an appropriate antigen as a vaccine protect animals from tumor growth in several animal models (Buschle et al., Gene Ther Mol Biol 1, (1998), 309-321; Schmidt et al., Proc Natl Acad Sci USA 94, (1997), 3262-3267). Thus, a vaccine consisting of polycationic compounds and antigen(s) is accepted in the art as being a very effective form of treatment.
GB 1 290 141 discloses a vaccine containing antigenic material with a base-rich peptide as an adjuvant. According to this document the vaccine may consist (as antigenic material) of live or killed organisms, whole or disrupted, or of preparations of natural toxins or products of the organism, or of preparations of extracts of the organisms, alone or in combination with one another. The base-rich peptide used as an adjuvant must contain at least 50% residues which have a free amino group, such as polylysine, polyornithine, polyarginine and polydiaminobutyric acids. In WO 97/30721 the use of basic polyamino acid as preferred adjuvant for vaccines in combination with an immunomodulating peptide or protein (fragment) is described. The use of a further immunostimulating substance to be used together with an antigen and such a polycationic polymer is neither disclosed nor made obvious by these two documents.
U.S. Pat. No. 3,725,545 describes that it is possible to potentiate the antibody production ability of nucleic acids containing preparations by adding cationically charged polymers in combination with single-stranded or multi-stranded nucleic acid polymers. Examples of such polycationic polymers are polyornithine, lysozyme, DEAE-Dextran, histone, hexadimethrine bromide and polylysine.
WO 91/04052 also relates to DEAE-Dextran as a polycationic adjuvant in a vaccine composition. This polycationic adjuvant is incorporated into a vaccine which further comprises the antigenic substance (the antigen) and saponin as a further adjuvant. Of course, neither polycationic compounds nor saponin as combined adjuvants may be regarded as an immunostimulating substance within the course of the present invention.